Fig. 5.

Examination of PTP1B function in EGF and PDGF signaling by differential SILAC labeling of PTP1B-deficient cells. A, Western blot analysis of tyrosine phosphorylated proteins in wild-type and KO cells under basal, EGF- (50 ng/ml, 5 min) or PDGF (20 ng/ml, 5 min)-stimulated conditions after 3 h of serum starvation. B, scheme depicting the strategy to analyze phosphorylation differences with quantitative MS upon EGF (50 ng/ml, 5 min) and PDGF (20 ng/ml, 5 min) stimulation in MEFs starved for 3 h. Each oval represents one independent SILAC experiment. Thereby not only alterations in phosphorylation between wild-type and KO cells can be examined, but ligand-induced phosphorylation events are also determined. pTyr, phosphotyrosine; wt, wild type.